Alterations of glomerular function have proven to be important in pathophysiological conditions such as diabetes mellitus (1,2). Glomerular hyperfiltration and hyperperfusion are characteristics of early diabetes in both human (1,3) and experimental animals (2,4). These glomerular hemodynamic alterations may play a role in the pathogenesis of the subsequent glomerular sclerosis associated with diabetes. The mechanisms mediating the increases in glomerular filtration of early diabetes, however, remain unknown. Recent observations on the mesangial cell indicate that these cells not only occupy a central anatomical position in the glomerulus, but also appear to play a central role in the regulation of glomerular function. Thus, a detailed examination of the changes provoked by diabetes mellitus on mesangial cell function is required to elucidate the mechanisms responsible for the initiation and progression of diabetic glomerulopathy. The overall objective of this proposal is to evaluate mesangial cell function in early diabetes. The specific aims are to: (1) establish and characterize an in vitro mesangial cell culture derived from streptozotocin-treated diabetic rats, and determine if the morphology of mesangial cells is altered in diabetes; (2) evaluate if prostaglandin synthesis and profile during basal and stimulated conditions is altered in "diabetic" mesangial cells; and (3) assess if the contractile response to vasoactive agents is impaired in "diabetic" mesangial cells and the correlation, if any, with alterations in prostaglandin synthesis, the mitogenic stage and/or changes in intracellular free calcium concentration. We will test the hypothesis that changes in mesangial cell function mediate the alterations in glomerular hemodynamics characteristic of the onset of diabetes mellitus. The basic experimental design will combine cell culture of mesangial cells obtained from normal and streptozotocin-diabetic rats with assessment of their physiological function (prostaglandin synthesis and contractility), cytoplasmic free calcium concentration, cell proliferative activity and morphological and morphometric determinations by light and conventional microscopy. The long term goal of the study is the parallel assessment of cell function, structure and proliferation, coupled with measurements of cytoplasmic free calcium ion levels, which will allow for the first time, an evaluation of mesangial cell function and structure during the pathogenesis of diabetic nephropathy. In addition, this research project will provide a unique training experience for both graduate and undergraduate students interested in learning and applying biomedical research techniques such as experimental animal surgery, cell culture methodology, light, fluorescence and electron microscopy, flow cytometry analysis, radioimmunoassay techniques, as well as statistical data analysis.